Osteoarthritis is the most common chronic condition of the joints. It is characterized by the degeneration of articular cartilage, formation of osteophytes and alterations in the synovium. This process has a severe impact on the quality of life of the patients and the currently available treatments are unsatisfactory and often merely focused on pain relief. In our group we are working on the development of in situ cross-linkable hydrogel platforms that could be used for resurfacing the damaged articular cartilage using a minimally invasive arthroscopic procedure. Stable fixation of the gel at the joint surface, facilitating the ingrowth of local stem and progenitor cell populations and supporting intrinsic repair mechanisms are considered minimal design parameters. To achieve this, we are exploring the use of enzymatically cross-linkable natural polymer-tyramine conjugates.

Dextran-tyramine conjugates were prepared by activation of dextran-OH and subsequent reaction with tyramine. Hyaluronic acid-tyramine and protein-tyramine conjugates were prepared using DMTMM coupling. In situ crosslinking is achieved by mixing the polymer conjugates with the enzyme HRP and minute, non-toxic amounts of H2O2 as oxidizing agent. Support of cartilage formation was studied after mixing of the polymer conjugates with mesenchymal stem cells, chondrocytes or combinations of both prior to crosslinking. Cell ingrowth was studied by implanting the hydrogels in an ex-vivo cartilage defect while mechanically loading the explant in a bioreactor and cell migration in the hydrogels was evaluated by tracking the sprouting of fluorescently labelled cell-spheroids.

We prepared dextran-tyramine conjugates with a degree of substitution of 10 tyramine residues per 100 monosaccharide units. The conjugated hyaluronic acid-tyramine had a degree of substitution of 10% of the carboxylic acid groups, while for the proteins the substitution was dependent on the protein type.

Enzymatically crosslinked hydrogels, based on dextran and hyaluronic acid, with the addition of co-cross linkable proteins show excellent properties for application in the regeneration of damaged cartilage.

Introduction: Local drug delivery yields higher gentamicin concentrations than can be safely achieved with systemic application. Unfortunately, both for beads as well as for bone cements, a sharp drop in release follows high initial gentamicin release. Aim of this study is to compare the effects of pulsed ultrasound on the release of gentamicin from antibiotic-loaded beads and bone cements. Mercury intrusion porosimetry is carried out to compare the pore size distribution in both materials before and after antibiotic release.

Materials and Methods: Ultrasound: Gentamicin release from three brands of gentamicin-loaded bone cement (CMW 1, Palamed G and Palacos R-G) and Septopal gentamicin-loaded beads was measured after 18 h of exposure in PBS to an ultrasonic field of 46.5 kHz in a 1:3 duty cycle with a peak intensity of 500 mW/cm² at the sample position. Ultrasound experiments were performed for 18h in 9-fold on bone cement and in 6-fold on beads. Samples not exposed to ultrasound were used as controls. The gentamicin release was measured with fluorescence polarisation immunoassay. Gentamicin release from insonated and control groups was compared using a two-tailed Student’s t test for independent samples.

Mercury intrusion porosimetry: In order to mimic bone cement and beads after prolonged stay in the human body (i.e. after initial release of the loaded gentamicin) samples were immersed for four and two weeks, in PBS. Immersed and not-immersed samples were compared.

Results: Pulsed ultrasound significantly enhanced gentamicin release from gentamicin-loaded beads, whereas gentamicin release from the gentamicin-loaded bone cements was not significantly enhanced. Mercury intrusion porosimetry revealed a rise in pores between 0.1 and 0.01 um in beads after gentamicin release, while in bone cements no increase in the number of pores before and after antibiotic release was found.

Conclusions: Ultrasound increases gentamicin release from antibiotic-loaded acrylic beads by 15%. Development of pores coincides with increased gentamicin release by ultrasound for beads. Application of ultrasound could optimise usage of an (antibiotic) reservoir in local drug delivery systems to treat bone and soft tissue infections more effectively.